Viruses invade all forms of life, causing diseases in humans including HIV-AIDS. Although the variety of viruses is daunting, all enveloped viruses, including HIV-1 and Rous Sarcoma Virus (RSV), must associate with the inner leaflet of plasma membranes. Our laboratories have undertaken a large-scale effort to characterize interactions between membrane components. The goal is to discover which membrane factors exert control over the interactions between membrane-associated viral matrix proteins. The overall approach is to combine experimental measurements, mainly FRET and fluorescence microscopy, with Monte Carlo computer simulations in order to discover all of the significant interaction energies between lipid-lipid, lipid-protein, and protein-protein. Membrane phase behavior and protein-protein interactions are first measured in model liposomes and then in live chicken fibroblast cells. The first specific aim is to develop the experiments and Monte Carlo simulations to describe gramicidin-gramicidin interactions in a simple 3-lipid component mixture containing cholesterol. The second aim is to handle the greatly increased complexity of a fifth component, ceramide or diacylglycerol. This aim includes proposed development of a "lipid buffer" in order to control the chemical potential of cholesterol, ceramide, or diacylglycerol in model or real biomembranes. The third aim is to characterize HIV-1 and RSV matrix protein-protein interactions in a lipid mixture that models the inner leaflet of an animal cell plasma membrane, as a basis for understanding behavior in live cells. The fourth and final aim is to examine HIV-1 and RSV matrix protein-protein interactions at the inner leaflet of the plasma membrane using FRET in live cells, attempting to find the factors that control the interactions, and whether coupling to the outer leaflet is observed. Useful treatments for viral afflictions, in particular HIV-AIDS, are not limited to "all or none" cures. Combinations of drugs, each of which slows down a step of the viral life cycle, have proven useful. The proposed research will lead to detailed understanding of how the membrane components cholesterol, ceramide, and diacylglycerol, all of which can be partially regulated by drugs, control one critical step in the life of the virus - association/assembly at the plasma membrane.